The genetic origins and diversity of Creole sheep from five regions of Colombia were investigated based on mitochondrial DNA (mtDNA) variations across 89 sequences from five breeds: one wool Creole sheep (CL) and four hair Creole sheep, including Ethiopian (OPCE), Sudan (OPCS), Pelibuey (OPCP) and Wayúu (OPCW). A global comparison was done using 62 haplotypes from Iberian, African, Indian, Caribbean, Mexican, Caucasian and European sheep based on sequences retrieved from GenBank. This study aimed to identify the maternal origin of Colombian Creole sheep and their genetic relationships at a global level. The results showed 31 different haplotypes from Colombian Creole sheep, which can be assigned to maternal lineage B, the most common lineage found in European sheep breeds and the only one found in several Iberian breed (e.g., Churra, Spanish Merino) that most likely participated in the Creole formation. Additional analyses showed that wool and hair sheep retained a broad genetic identity despite being geographically separated. The global-level phylogenetic analysis revealed that Colombian Creole sheep belong to a distinct and defined genetic lineage that is likely the result of a founder effect with ecotypes of Iberian descent and the subsequent introduction of foreign breeds. This is consistent with historical reports on the presence of sheep in South America and, particularly, Colombia.